ANTI-T CELL IMMUNOTOXINS - A LOOK AT POST-ENDOCYTOTIC RECEPTOR-MEDIATED ROUTING

Many types of targeted drug delivery systems utilize cell surface rece ptor binding to achieve targeting specificity. Receptor-mediated endoc ytosis also provides internalization of the targeted drug. What has no t been previously appreciated is the fact that cell surface receptors play an important post-endocytotic role. After endocytosis the recepto r ligand interaction determines how the ligand-drug complex will be ro uted. In many cases this routing uniquely determines the overall effic acy of the drug delivery system. In this paper we will show that a uni que anti-T cell immunotoxin, anti-CD3+-CRM9, constructed with a diphth eria toxin binding site mutant, CRM9, is a highly effective reagent fo r inducing in vivo T cell ablation. The high degree of efficacy and th e wide therapeutic margin of this reagent is a consequence of the obli gatory intracellular routing pathway required for diphtheria toxin (DT ) intoxication. The DT receptor (DTR) typically fulfills this function in diphtheria toxin intoxication. In the case of the anti-CD3+ toxin conjugate the routing function is performed by CD3. This epitope route s parallel with DTR. This fact accounts for the high toxicity of anti- CD3 conjugates made with diphtheria toxin binding site mutants such as CRM9. CRM9 a one is incapable of entering the optimal routing pathway and hence has very low systemic toxicity. The anti-CD3 antibody doubl y complements the CRM9 defect: It facilitates ( 1 ) the entrance of CR M9 into the targeted cell endosomes and (2 directs the entrance of CRM 9 into the DT intoxication pathway. It is likely that these concepts w ill have relevance to other cell type-specific immunotoxins and other cell type-specific targeted delivery systems which depend on the membr ane translocation of bio-active macromolecules into the cytosol or nuc lear compartments.